Grain size is one of the most important factors that control rice yield,as it is associated with grain weight(GW).To date,dozens of rice genes that regulate grain size have been isolated;however,the regulatory mechani...Grain size is one of the most important factors that control rice yield,as it is associated with grain weight(GW).To date,dozens of rice genes that regulate grain size have been isolated;however,the regulatory mechanism underlying GW control is not fully understood.Here,the quantitative trait locus qGL5 for grain length(GL)and GW was identified in recombinant inbred lines of 9311 and Nipponbare(NPB)and fine mapped to a candidate gene,OsAUX3.Sequence variations between 9311 and NPB in the OsAUX3 promoter and loss of function of OsAUX3 led to higher GL and GW.RNA sequencing,gene expression quantification,dual-luciferase reporter assays,chromatin immunoprecipitation-quantitative PCR,and yeast one-hybrid assays demonstrated that OsARF6 is an upstream transcription factor regulating the expression of OsAUX3.OsARF6 binds directly to the auxin response elements of the OsAUX3 promoter,covering a single-nucleotide polymorphism site between 9311 and NPB/Dongjin/Hwayoung,and thereby controls GL by altering longitudinal expansion and auxin distribution/content in glume cells.Furthermore,we showed that miR167a positively regulate GL and GW by directing OsARF6 mRNA silencing.Taken together,our study reveals that a novel miR167a-OsARF6-OsAUX3 module regulates GL and GW in rice,providing a potential target for the improvement of rice yield.展开更多
Argonaute proteins(AGOs) are central to RNA silencing pathways and play critical roles in plant antiviral defense.However,the functions of individual AGOs in rice remain incompletely understood.In this study,we demons...Argonaute proteins(AGOs) are central to RNA silencing pathways and play critical roles in plant antiviral defense.However,the functions of individual AGOs in rice remain incompletely understood.In this study,we demonstrate that rice AGO2contributes to resistance against rice ragged stunt virus(RRSV) through a regulatory module involving mi R167g-3p and its target gene,SNAP32.Immunoprecipitation coupled with small RNA sequencing revealed that AGO2 associates not only with virus-derived small interfering RNAs(vsi RNAs)but also preferentially associates with mi R167g-3p during RRSV infection.Functional analyses further showed that mi R167g-3p expression is induced upon infection.Transgenic rice lines overexpressing mi R167g-3p exhibited enhanced resistance,whereas knockdown lines were more susceptible.SNAP32 was validated as a direct target of mi R167g-3p through transient expression assays in Nicotiana benthamiana and dual-luciferase assays in rice protoplasts.Expression analyses confirmed that mi R167g-3p represses SNAP32 at the transcript level.Consistently,SNAP32-overexpressing plants displayed increased susceptibility to RRSV,while snap32 knockout plants showed enhanced resistance,supporting a negative role of SNAP32 in antiviral defense.Together,these findings establish a regulatory pathway in which AGO2 promotes antiviral immunity by stabilizing mi R167g-3p to repress SNAP32,thereby restricting RRSV infection.This work advances our understanding of AGO2-mediated defense in rice and highlights the use of a mi RNA 3p strand within an AGO-mi RNA-target module as an important layer of resistance against viral pathogens.展开更多
Grain weight and quality are always determined by grain filling.Plant microRNAs have drawn attention as key targets for regulation of grain size and yield.However,the mechanisms that underlie grain size regulation rem...Grain weight and quality are always determined by grain filling.Plant microRNAs have drawn attention as key targets for regulation of grain size and yield.However,the mechanisms that underlie grain size regulation remain largely unclear because of the complex networks that control this trait.Our earlier studies demonstrated that suppressed expression of miR167(STTM/MIM167)substantially increased grain weight.In a field test,the yield increased up to 12.90%-21.94% because of a significantly enhanced grain filling rate.Here,biochemical and genetic analyses revealed the regulatory effects of miR159 on miR167 expression.Further analysis indicated that OsARF12 is the major mediator by which miR167 regulates rice grain filling.Overexpression of OsARF12 produced grain weight and grain filling phenotypes resembling those of STTM/MIM167 plants.Upon in-depth analysis,we found that OsARF12 activates OsCDKF;2 expression by directly binding to the TGTCGG motif in its promoter region.Flow cytometry analysis of young panicles from OsARF12-overexpressing plants and examination of cell number in cdkf;2 mutants verified that OsARF12 positively regulates grain filling and grain size by targeting OsCDKF;2.Moreover,RNA sequencing results suggested that the miR167-OsARF12 module is involved in the cell development process and hormone pathways.OsARF12-overexpressing plants and cdkf;2 mutants exhibited enhanced and reduced sensitivity to exogenous auxin and brassinosteroid(BR)treatment,confirming that targeting of OsCDKF;2 by OsARF12 mediates auxin and BR signaling.Our results reveal that the miR167-OsARF12 module works downstream of miR159 to regulate rice grain filling and grain size via OsCDKF;2 by controlling cell division and mediating auxin and BR signals.展开更多
MicroRNAs(miRNAs)are a class of small(20–24 nucleotides(nt)long)non-coding RNAs.One mature miRNA can be transcribed from one or more gene loci known as miRNA genes(MIRs).The transcript of a MIR forms a stem-loop stru...MicroRNAs(miRNAs)are a class of small(20–24 nucleotides(nt)long)non-coding RNAs.One mature miRNA can be transcribed from one or more gene loci known as miRNA genes(MIRs).The transcript of a MIR forms a stem-loop structure that is processed into a 20–24-nt miRNA-5p/−3p duplex by RNase III family endoribonucleases such as Dicer-like1(DCL1).In turn,the overhang ends of the duplex are methylated by HUA ENHANCER 1(HEN1),generating stabilized mature miRNAs.The mature miRNAs are loaded onto ARGONAUTE(AGO)proteins,forming a miRNAinduced gene silencing complex(miRISC).Then,the miRISC binds to target sites with sequences complementary to the miRNAs,leading to either cleavage or translational inhibition of the target mRNAs,or methylation of the target sequences,resulting in post-transcriptional and transcriptional gene silencing,respectively.In the past decade,more than 700 miRNAs have been identified in rice,a subset of which have been found to be responsive to the rice blast fungus,Magnaporthe oryzae,or its elicitors.Moreover,members of 10 miRNA families have been found to positively or negatively regulate rice defense against M.oryzae,namely miR160,miR164,miR166,miR167,miR169,miR319,miR396,miR398,miR444 and miR7695.This review summarizes the identification and functional characterization of the miRNAs,which respond to M.oryzae or its elicitors and describes the current understanding of the complicated but wellorganized network in the context of rice-M.oryzae interaction.展开更多
基金This project was funded by grants from the National Key Research and Development Program of China(2016YFD0100400)the National Natural Science Foundation of China(32060451)the Zhejiang Provincial Nat-ural Science Foundation of China(grant no.L Z19C020001).
文摘Grain size is one of the most important factors that control rice yield,as it is associated with grain weight(GW).To date,dozens of rice genes that regulate grain size have been isolated;however,the regulatory mechanism underlying GW control is not fully understood.Here,the quantitative trait locus qGL5 for grain length(GL)and GW was identified in recombinant inbred lines of 9311 and Nipponbare(NPB)and fine mapped to a candidate gene,OsAUX3.Sequence variations between 9311 and NPB in the OsAUX3 promoter and loss of function of OsAUX3 led to higher GL and GW.RNA sequencing,gene expression quantification,dual-luciferase reporter assays,chromatin immunoprecipitation-quantitative PCR,and yeast one-hybrid assays demonstrated that OsARF6 is an upstream transcription factor regulating the expression of OsAUX3.OsARF6 binds directly to the auxin response elements of the OsAUX3 promoter,covering a single-nucleotide polymorphism site between 9311 and NPB/Dongjin/Hwayoung,and thereby controls GL by altering longitudinal expansion and auxin distribution/content in glume cells.Furthermore,we showed that miR167a positively regulate GL and GW by directing OsARF6 mRNA silencing.Taken together,our study reveals that a novel miR167a-OsARF6-OsAUX3 module regulates GL and GW in rice,providing a potential target for the improvement of rice yield.
基金supported by grants from the Biological Breeding-National Science and Technology Major Project(2023ZD04070)the National Natural Science Foundation of China (Nos. 32025031,31871934,32170167)the Natural Science Foundation of Fujian Province (2025J01549)。
文摘Argonaute proteins(AGOs) are central to RNA silencing pathways and play critical roles in plant antiviral defense.However,the functions of individual AGOs in rice remain incompletely understood.In this study,we demonstrate that rice AGO2contributes to resistance against rice ragged stunt virus(RRSV) through a regulatory module involving mi R167g-3p and its target gene,SNAP32.Immunoprecipitation coupled with small RNA sequencing revealed that AGO2 associates not only with virus-derived small interfering RNAs(vsi RNAs)but also preferentially associates with mi R167g-3p during RRSV infection.Functional analyses further showed that mi R167g-3p expression is induced upon infection.Transgenic rice lines overexpressing mi R167g-3p exhibited enhanced resistance,whereas knockdown lines were more susceptible.SNAP32 was validated as a direct target of mi R167g-3p through transient expression assays in Nicotiana benthamiana and dual-luciferase assays in rice protoplasts.Expression analyses confirmed that mi R167g-3p represses SNAP32 at the transcript level.Consistently,SNAP32-overexpressing plants displayed increased susceptibility to RRSV,while snap32 knockout plants showed enhanced resistance,supporting a negative role of SNAP32 in antiviral defense.Together,these findings establish a regulatory pathway in which AGO2 promotes antiviral immunity by stabilizing mi R167g-3p to repress SNAP32,thereby restricting RRSV infection.This work advances our understanding of AGO2-mediated defense in rice and highlights the use of a mi RNA 3p strand within an AGO-mi RNA-target module as an important layer of resistance against viral pathogens.
基金funded by the National Natural Science Foundation of China(NSFC,32272014,32001440,31971846,and 31871554)the Natural Science Foundation of Henan Province-Excellent Youth Fund(222300420049)+2 种基金the Central Plains Talents Program of Henan Province(Talent Training Series)-Top Young Talents in Central Plains(ZYY-CYU202012170)the Support Plan for Scientific and Technological Innovation Talents in Colleges and Universities of Henan Province(21HAS-TIT037)the China Postdoctoral Science Foundation(2020M682294).
文摘Grain weight and quality are always determined by grain filling.Plant microRNAs have drawn attention as key targets for regulation of grain size and yield.However,the mechanisms that underlie grain size regulation remain largely unclear because of the complex networks that control this trait.Our earlier studies demonstrated that suppressed expression of miR167(STTM/MIM167)substantially increased grain weight.In a field test,the yield increased up to 12.90%-21.94% because of a significantly enhanced grain filling rate.Here,biochemical and genetic analyses revealed the regulatory effects of miR159 on miR167 expression.Further analysis indicated that OsARF12 is the major mediator by which miR167 regulates rice grain filling.Overexpression of OsARF12 produced grain weight and grain filling phenotypes resembling those of STTM/MIM167 plants.Upon in-depth analysis,we found that OsARF12 activates OsCDKF;2 expression by directly binding to the TGTCGG motif in its promoter region.Flow cytometry analysis of young panicles from OsARF12-overexpressing plants and examination of cell number in cdkf;2 mutants verified that OsARF12 positively regulates grain filling and grain size by targeting OsCDKF;2.Moreover,RNA sequencing results suggested that the miR167-OsARF12 module is involved in the cell development process and hormone pathways.OsARF12-overexpressing plants and cdkf;2 mutants exhibited enhanced and reduced sensitivity to exogenous auxin and brassinosteroid(BR)treatment,confirming that targeting of OsCDKF;2 by OsARF12 mediates auxin and BR signaling.Our results reveal that the miR167-OsARF12 module works downstream of miR159 to regulate rice grain filling and grain size via OsCDKF;2 by controlling cell division and mediating auxin and BR signals.
基金supported by the National Natural Science Foundation of China grants(31430072 and 31672090 to W-MW,and 31471761 to YL).
文摘MicroRNAs(miRNAs)are a class of small(20–24 nucleotides(nt)long)non-coding RNAs.One mature miRNA can be transcribed from one or more gene loci known as miRNA genes(MIRs).The transcript of a MIR forms a stem-loop structure that is processed into a 20–24-nt miRNA-5p/−3p duplex by RNase III family endoribonucleases such as Dicer-like1(DCL1).In turn,the overhang ends of the duplex are methylated by HUA ENHANCER 1(HEN1),generating stabilized mature miRNAs.The mature miRNAs are loaded onto ARGONAUTE(AGO)proteins,forming a miRNAinduced gene silencing complex(miRISC).Then,the miRISC binds to target sites with sequences complementary to the miRNAs,leading to either cleavage or translational inhibition of the target mRNAs,or methylation of the target sequences,resulting in post-transcriptional and transcriptional gene silencing,respectively.In the past decade,more than 700 miRNAs have been identified in rice,a subset of which have been found to be responsive to the rice blast fungus,Magnaporthe oryzae,or its elicitors.Moreover,members of 10 miRNA families have been found to positively or negatively regulate rice defense against M.oryzae,namely miR160,miR164,miR166,miR167,miR169,miR319,miR396,miR398,miR444 and miR7695.This review summarizes the identification and functional characterization of the miRNAs,which respond to M.oryzae or its elicitors and describes the current understanding of the complicated but wellorganized network in the context of rice-M.oryzae interaction.
